Increasing the amylose content of maize through silencing of sbe2a genes
Improved amylose content in maize has been achieved by reducing the starch branching enzyme (SBE) activity via transgenic maize inbred line transformed by a high-efficient RNAi expression vector, which may provide the foundation for maize quality improvement. The sense and anti-sense fragments of maize SBE gene sbe2a were cloned by reverse transcript PCR and high efficient RNAi expression vector was constructed based on plant expression plasmid pCAMBIA1301. Then the reconstruct was introduced into maize inbred line Tie7922 by pollen tube pathway transformation. Four transgenic plants were obtained. The integration of interest gene sbe2a into maize genome has been confirmed by PCR amplification and Southern hybridization. The sbe2a transcription was suppressed obviously by the analyses of RT-PCR, SBE activity and amylase content on the four transgenic plants. The SBE activity was significantly less than that of wild type maize, and was at most reduced by 77.9%; the amylose content was at most increased by 87.8%. In conclusion, RNAi expression vector pRSBE2a containing sbe2a gene was successfully constructed. Through genetic transformation, RNAi technique efficiently silences endogenous sbe2a gene to reduce the SBE activity, and high-amylose maize linesare obtained.
Keywords: Maize, starch branching enzyme gene sbe2a, RNA interference, genetic transformation